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1. (WO2018228071) PREPARATION AND AMPLIFICATION CULTURE METHODS FOR HUMAN PLURIPOTENT STEM-CELL-DERIVED HUMAN RETINAL PIGMENT EPITHELIAL CELL
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Pub. No.: WO/2018/228071 International Application No.: PCT/CN2018/085026
Publication Date: 20.12.2018 International Filing Date: 28.04.2018
IPC:
C12N 5/079 (2010.01) ,C12N 5/0735 (2010.01) ,C12N 5/071 (2010.01)
C CHEMISTRY; METALLURGY
12
BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
N
MICRO-ORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICRO-ORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
5
Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
07
Animal cells or tissues
071
Vertebrate cells or tissues, e.g. human cells or tissues
079
Neural cells
C CHEMISTRY; METALLURGY
12
BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
N
MICRO-ORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICRO-ORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
5
Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
07
Animal cells or tissues
071
Vertebrate cells or tissues, e.g. human cells or tissues
073
Embryonic cells or tissues; Foetal cells or tissues
0735
Embryonic stem cells; Embryonic germ cells
C CHEMISTRY; METALLURGY
12
BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
N
MICRO-ORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICRO-ORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
5
Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
07
Animal cells or tissues
071
Vertebrate cells or tissues, e.g. human cells or tissues
Applicants:
中山大学中山眼科中心 ZHONGSHAN OPHTHALMIC CENTER, SUN YAT-SEN UNIVERSITY [CN/CN]; 中国广东省广州市 越秀区先烈南路54号 No.54, Xianlie South Road, Yuexiu District Guangzhou, Guangdong 510060, CN
Inventors:
钟秀风 ZHONG, Xiufeng; CN
葛坚 GE, Jian; CN
刘胜旭 LIU, Shengxu; CN
彭福华 PENG, Fuhua; CN
Agent:
广州科粤专利商标代理有限公司 GUANGZHOU KEYUE I.P. LAW OFFICE; 中国广东省广州市 越秀区先烈中路100号大院23-1栋616室 Room 616, 6/F, Building 23-1 No.100, Xianlie Central Road, Yuexiu District Guangzhou, Guangdong 510070, CN
Priority Data:
201710441265.913.06.2017CN
Title (EN) PREPARATION AND AMPLIFICATION CULTURE METHODS FOR HUMAN PLURIPOTENT STEM-CELL-DERIVED HUMAN RETINAL PIGMENT EPITHELIAL CELL
(FR) PROCÉDÉS DE PRÉPARATION ET DE CULTURE PAR AMPLIFICATION POUR CELLULE ÉPITHÉLIALE DE PIGMENT RÉTINIEN HUMAIN DÉRIVÉE DE CELLULES SOUCHES PLURIPOTENTES HUMAINES
(ZH) 一种人多能干细胞来源人视网膜色素上皮细胞的制备和扩增培养方法
Abstract:
(EN) Provided are preparation and amplification culture methods for a human pluripotent stem-cell-derived human retinal pigment epithelial cell. The preparation method comprises the following steps of: collecting human pluripotent stem-cell-derived 3D-RPE spheres, mechanically isolating and removing pigment-free non-RPE cells or mass, leaving pigment-containing RPE cell debris; enzymatically hydrolyzing and digesting the pigment-containing RPE cell debris, obtaining a RPE single cell suspension, and thereby obtaining the human pluripotent stem-cell-derived human retinal pigment epithelial cell. The amplification method comprises centrifuging the RPE single cell suspension followed by discarding the supernatant and then resuspending same with the RPE cell culture medium, inoculating the cells onto the cell culture container pre-coated with an extracellular matrix for primary culture, and carrying out subculture after the cells completely fill the container.
(FR) L'invention concerne des procédés de préparation et de culture par amplification pour une cellule épithéliale de pigment rétinien humain dérivée de cellules souches pluripotentes humaines. Le procédé de préparation comprend les étapes suivantes : collecte de sphères de RPE 3D dérivées de cellules souches pluripotentes humaines, isolement et élimination mécanique de cellules non RPE exemptes de pigment ou d'une masse, laissant des débris de cellules RPE contenant un pigment; hydrolyse enzymatique et digestion des débris de cellules RPE contenant un pigment, obtention d'une suspension de cellules uniques RPE, et ainsi obtention de la cellule épithéliale de pigment rétinien humain dérivée de cellules souches pluripotentes humaines. Le procédé d'amplification comprend la centrifugation de la suspension cellulaire unique RPE suivie de l'élimination du surnageant puis sa remise en suspension avec le milieu de culture de cellule RPE, l'inoculation des cellules sur le récipient de culture cellulaire pré-revêtu d'une matrice extracellulaire pour la culture primaire, et la réalisation d'une sous-culture après que les cellules ont complètement rempli le récipient.
(ZH) 提供了人多能干细胞来源人视网膜色素上皮细胞的制备和扩增培养方法。该制备方法包括以下步骤:收集人多能干细胞来源的3D-RPE球体,机械分离、去除不含色素的非RPE细胞或团块,留取含有色素的RPE细胞片;酶解消化含有色素的RPE细胞片,获得RPE单细胞悬液,由此获得人多能干细胞来源人视网膜色素上皮细胞。该扩增方法包括将RPE单细胞悬液离心后弃上清再用RPE细胞培养基重悬,接种到预先包被了细胞外基质的细胞培养容器中进行原代培养,细胞长满后进行传代培养。
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Designated States: AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IR, IS, JO, JP, KE, KG, KH, KN, KP, KR, KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW
African Regional Intellectual Property Organization (ARIPO) (BW, GH, GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, TZ, UG, ZM, ZW)
Eurasian Patent Office (AM, AZ, BY, KG, KZ, RU, TJ, TM)
European Patent Office (EPO) (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, IE, IS, IT, LT, LU, LV, MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, TR)
African Intellectual Property Organization (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, KM, ML, MR, NE, SN, TD, TG)
Publication Language: Chinese (ZH)
Filing Language: Chinese (ZH)